The staphylococcal enterotoxins are extracellular proteins composed of sinule polypeptide chains of about 30 kd that characteristically have a disulfide loop near the middle of the molecule. They are categorized into five serological groups, designated Staphylococcal enterotoxin A (SEA), Staphylococcal enterotoxin B (SEB), Staphylococcal enterotoxin C (SEC), Staphylococcal enterotoxin D (SED) and Staphylococcal enterotoxin E (SEE). Based on differences in minor epitopes, SEC has been further subdivided into three types designated SEC.sub.1, SEC.sub.2 and SEC.sub.3. A sixth group, SEF, was also described and was implicated as the causative agent of toxic shock syndrome. It has since been discounted as an enterotoxin and renamed toxic shock syndrome toxin-1 (TSST-1), as cited by J. J. Iandolo in Ann. Rev. of Micro. Vol. 43, pp. 275-402, 1989.
Menstrually occurring toxic shock syndrome is a severe and sometimes fatal multi-system disease associated with infection or colonization by Staphylococcus aureus (S. aureus) bacteria, has been linked to the use of tampons during menstruation. In Toxic shock syndrome(TSS), whether associated with menstruation or not, the symptoms include fever, hypotension, rash, and desquamation of the skin. TSST-1 is highly associated with menstrual cases but is less often isolated from Staphylococcus aureus strains in non-menstrual cases of the illness. Since TSST-1 can induce many clinical features of TSS in the rabbit and other species, it is generally thought to be the causative toxin in TSS (Schlievert, "Staphylococcal Enterotoxin B and Toxic Shock Syndrome Toxin-1 Are Significantly Associated With Non-menstrual TSS", The Lancet, Vol. 1(8490), May 17, 1986, p.1149). However, Garbe (Garbe P. L., Arko R. J., Reingold A. L., et al. "Staphylococcus aureus isolates from patients with non-menstrual toxic shock syndrome: Evidence for additional toxins", JAMA 1985; Vol 253; pp. 2538-42) noted that many TSS isolates from nonmenstrual cases did not express TSST-1 though they did cause TSS-like symptoms in a rabbit model. Of the toxins formed by S. aureus nonmenstrual isolates, TSST-1 was produced by 40% of those reported by Schlievert, 1986. Further, enterotoxin B was made by 38% of all non-menstrual TSS strains. Furthermore, Schlievert reported about 78% of non-menstrual TSS or probable TSS isolates expressed either TSST-1 or enterotoxin B compared with 20% of non-menstrual non-TSS isolates (p&lt;0.001).
Isolates from thirty patients suffering from confirmed or probable toxic shock syndrome were also examined for enterotoxins and exfoliative toxins. It was found that enterotoxin B was made by 38% of all nonmenstrual TSS strains. Crass and Bergdoll, 1986 reported a total of 46 (83.6%) of 55 S. aureus isolates produced TSST-1: 12 (25.5%) alone, 21 (46.8%) with Staphylococcal Enterotoxin A and 13 (27.7%) with Staphylococcal Enterotoxin C. Eight of the S. aureus isolates that did not produce TSST-1 did produce Staphylococcal enterotoxin B.
Humphreys (1989) reported the studies of S. aureus isolates obtained from cases of septicaemia, in which 33 (63%) produced enterotoxins A, B, C, or D alone or in combination.
Toxic shock syndrome toxin 1 (TSST-1) is classified as a member of the pyrogenic exotoxin-staphylococcal enterotoxin group of toxins on the basis of a patient's symptoms. However, TSST-1 and its cognate antiserum do not cross-react with any of the sera or proteins of other members of this toxin family. Furthermore, TSST-1 lacks the cysteine loop present in the remaining members of the toxin family, an important structural feature of this family of toxins. TSST-1 has very little amino acid homology with other members of this toxin family. The lack of a close sequence relationship between TSST-1 and other toxins may suggest that it is more closely related to the ancestral progenitor of this family or, alternatively, is wrongly included in this group of toxins (Iandolo, 1989). Nevertheless, TSST-1 bears little structural relationship to the other members of the pyrogenic exotoxin-staphylococcal enterotoxin group (Iandolo, 1989).
Subsequent to the publication of reports associating toxic shock syndrome with the use of tampons, a number of investigators undertook studies designed to evaluate the effect of tampons on growth of S. aureus bacteria as well as the effect of tampons on the production of TSST-1 by that bacteria. Early efforts to elucidate the role of tampons in TSS yielded conflicting data. Schlievert et al. (Obstet. Gynecol., Vol. 64, pp. 666-670, November 1984) studied the effect of tampons on S. aureus to evaluate whether or not tampon components increase growth of S. aureus and production of toxic shock syndrome toxin-1. It was concluded that, under the test conditions of their study, tampon components provide neither nutrients for growth of toxic shock syndrome S. aureus nor factors that induce production of toxic shock syndrome toxin-1 above control levels. After six-hours' incubation, some commercially available tampons which were tested were inhibitory to bacterial growth and suppressed toxin production. Others suppressed toxin production but did not inhibit cell growth. One tampon inhibited cell growth but increased the amount of toxin produced. On the other hand, Tierno and Hanna (Contraception, Vol. 31, pp 185-194, 1985) reported that in their experiments tampons did stimulate S. aureus to produce TSST-1.
Reiser et al. (J. Clin. Microbiol., Vol. 25, No. 8, pp. 1450-1452, August 1987) thereafter reported the results of tests they conducted to determine the effect of four brands of tampons on production of toxic shock syndrome toxin-1. The amount of air available to the tampons which were tested was limited to that contained in sacs (made from cellulose sausage casing with a molecular weight cut-off of less than 10,000) in which the tampons were enclosed during testing. This method was deemed advantageous in that the limited amount of available air was thought to mimic more closely than previously used methods the in vivo condition in the vagina during menstruation with a tampon in place and in that the tampons which were tested were not altered prior to testing. The results of the tests conducted by Reiser et al. indicated that tampons provide increased surface area for the S. aureus bacteria to grow and adequate oxygen for toxin production. No significant inhibition of growth of the staphylococci bacteria or TSST-1 production by any of the tampons tested was noted.
Robbins et al., publishing in J. Clinical Microbiol., Vol. 25, No. 8, pp. 1446-1449, August 1987 at the same time as Reiser et al., reported the effect of 17 commercially available tampons on TSST-1 toxin production using a disk-membrane-agar (DMA) method, with incubation at 37.degree. C. for 19 hours under 5% CO.sub.2 in air. Filter membranes overlaying agar medium (with or without blood) in small petri dishes were spread inoculated with a TSST-1 producing strain of S. aureus. Robbins et al. concluded that the main role of tampons in TSS may be that of providing a fibrous surface for heavy colonization and sufficient air for TSST-1 production. In addition, they found evidence of inhibition of TSST-1 production by additives such as the deodorant/surfactant used in a commercially available deodorant tampon and a decrease in TSST-1 production by inhibiting growth of S. aureus as was observed in the case of a different commercially available tampon. It was thought that both inhibition of TSST-1 production and inhibition of S. aureus growth might prove to be important in reducing the risk of TSS.
U.S. Pat. No. 4,405,323 to Auerbach discloses a tampon designed to eliminate the hazards of toxic shock syndrome and dysmenorrhea. The tampon has incorporated therein an antibacterial agent which is said to disperse on contact with body fluids and prevent development of the organisms which produce the toxins which cause toxic shock syndrome. Among the antibacterial materials disclosed for use are povidone-iodine compound, mercury, zinc, penicillin, erythromycin and nitrofurazone.
Patent Cooperation Treaty Publication No. WO 86/05388 (published Sept. 25, 1986) to Kass teaches that the inclusion of a salt of a nontoxic divalent cation in absorptive pads, e.g. catemenial tampons, inhibits production of toxic shock syndrome toxin-1 and other staphylococcal products during use of said absorptive pad. Suitable salts include those of magnesium, barium, calcium or strontium (preferred) or of other divalent cations such as zinc, manganese, copper, iron, nickel and the like. The anionic portion of the salt is not critical. Magnesium stearate and magnesium acetate are particularly preferred salts for use in the invention.
U.S. Pat. No. 4,374,522 to Olevsky states that patterns of use of catemenial tampon seem to indicate that high absorptive capacity with the concomitant extended period of use of certain tampons are factors which contribute to the formation of toxic shock syndrome. The invention theorizes that tampons having limited absorptive capacity and requiring relatively more frequent changes may be desirable. The Olevsky patent provides a tampon made of conventional cellulosic materials, such as rayon fibers, which have been compressed into a bullet-shape with an open bottom surface sealed by a fluid impermeable sheet. The fluid impermeable bottom and the traditional bullet shaped pledget define a hollow core central reservoir area which is said to serve as a reservoir for excess menstrual fluid.
U.S. Pat. No. 4,431,427 to Lefren et al. discloses menstrual tampons comprising physiologically safe, water-soluble acids in their monomeric, oligomeric or polymeric forms. Citric, glycolic, malic, tartaric and lactic acids are disclosed as being useful in the practice of the invention. The presence of one or more of the above-noted acids in a tampon is said to inhibit the growth of bacteria responsible for toxic shock. Where an acid is used in its polymeric form, the tampon may additionally include an enzyme to hydrolyze the polymeric acid to its monomeric form.
Canadian Patent No. 1,123,155 to Sipos discloses a catemenial tampon for preventing toxic shock syndrome during menstrual flow. The body of the tampon, which is open at the insertion end and is closed at the withdrawal end, is snugly surrounded in its expanded condition by a fluid proof, thin and flexible membrane. This membrane, which can be made of polyethylene sheet, is biased against the vaginal wall during use of the tampon, is neutral to the vaginal mucosa and is completely impermeable to bacteria, viruses and toxic decomposition products of the menstrual flow.
Canadian Patent No. 1,192,701 to Bardhan discloses a tampon for the absorption of menstrual flow and comprising an inner layer of liquid-absorbent material and an outer layer which surrounds and encloses the inner layer. Menstrual discharge may flow inwardly to the inner layer but the outer layer is impervious to the passage of menstrual fluid outwardly from the inner layer. A plurality of liquid absorbent wicks extending from the inner layer through apertures formed in the outer layer serve as conduits for the flow of menstrual discharge from outside the tampon to the inner layer thereof. The disclosed structure is said to minimize the availability of discharge outside the tampon with a resulting reduction in the likelihood of growth of S. aureus and consequently its production of toxin. This patent also discloses that an antimicrobial compound which is bactericidal or bacteriostatic to S. aureus may be included in the inner layer. The antimicrobial agent may take the form of an antibiotic (such as penicillin, erythromycin, tetracycline or neomycin), a chemotherapeutic agent (such as a sulfonamide) or a disinfectant (such as phenol). The patent states that because the tampon is protected by its outer layer from contact with the vaginal wall, the risk of an allergic or other adverse reaction to the anti-microbial agent is minimized, and since the antimicrobial agent is also protected by the outer layer from contact with menstrual discharge, there is little risk of the destruction of commensal organisms in the vagina or development of resistance to the antimicrobial agent by S. aureus in any menstrual discharge outside the vagina.
S. Notermans et al. (Journal of Food Safety, Vol. 3 (1981), pages 83-88) reported that glyceryl monolaurate, when used in the proportion of 5g per kg. of meat slurry (pH 6.0-6.2) inhibited toxin productions by Clostridium botulinum type A, type B and type E. This article does not mention Staphylococcus aureus nor any toxins produced therefrom nor does it mention absorbent products or toxic shock syndrome.
U.S. Pat. No. 4,585,792 to Jacob et al. discloses that L-ascorbic acid when topically applied to the vaginal area of a human female during menses will inactivate toxins known to contribute to Toxic Shock Syndrome. The ascorbic acid compound may be carried by a vaginal tampon. The disclosure of U.S. Pat. No. 4,722,937, is to the same effect.
U.S. Pat. No. 4,413,986 to Jacobs discloses a tampon assembly packaged for sterile insertion of a tampon into the vagina having a guide tube telescoped around an insertion tube and a flexible sheath attached to the inner end of the guide tube and tucked into the inner end of the insertion tube. In use, as the insertion tube is pushed through the guide tube and into the vagina, the flexible sheath is pulled over the inner end of the insertion tube and extends along the exterior thereof. The portion of the insertion tube which is inserted into the vagina is at all times fully sheathed by the flexible sheath.
In tampon-associated toxic shock syndrome cases, the predominant toxin produced by S. aureus is toxic shock syndrome toxin-1 (TSST-1), while to a lesser extent, other enterotoxins can be produced. These non-TSST-1 enterotoxins are principally associated with nonmenstrual toxic shock syndrome.
There are compounds known to affect enterotoxin production. J. L. Smith, M. M. Bencievengo, R. L. Buchanan, and C. A. Kunsch reported, in an article entitled "Effect of Glucose Analogs on the Synthesis of Staphylococcal Enterotoxin A", Journal of Food Safety 8 (198) pp. 139-146 that glucose, 2-deoxyglucose and alpha-methyl glucose inhibited staphylococcal enterotoxin A synthesis by Staphylococcus aureus 196E, whereas beta-methyl glucose and 3-0-methyl glucose did not inhibit synthesis even at high concentrations. The formation of enterotoxins A and B is inhibited by glucose, with the extent of inhibition being strongly influenced by pregrowth of the bacteria in a glucose-containing medium as cited by Smith et al.
Iandolo and Shafer reported the effect of glucose and glucose analogs 2-deoxyglucose and alpha methyl glucoside on the synthesis and regulation of Staphylococcal enterotoxin B production (Iandolo and Shafer, "Regulation of Staphylococcal Enterotoxin B", Infection and Immunity, May 1977, pp. 610-615). The attenuating effect of glucose on Staphylococcal enterotoxin B was observed. However, when this effect was examined with analogs of glucose, contradictory responses were seen. Acid production due to glucose metabolism has been shown to reduce toxin production.
Ibrahim, Radford, Baldock and Ireland reported data indicating that, in cheese without starter activity, inhibition of growth of S. aureus and enterotoxin production may be achieved during production by not salting the curd at the end of cheddaring, avoiding pressing at high ambient temperatures and minimizing the pressing time of the curd (Ibrahim, et al., "Inhibition of Growth of Staphylococcus aureus and Enterotoxin-A Production in Cheddar Cheese Produced with Induced Starter Failure", Journal of Food Protection, Vol. 44, No. 3, pp. 189-193, March 1981). The authors also reported that storage of salted cheese at 11.degree. C. appears to be a potential hazard because of significant increases in S. aureus count and entertoxin concentration. Surprisingly, the authors reported that the count of S. aureus decreased with no change in enterotoxin concentration in unsalted cheese stored at 11.degree. C.
Another study was reported by J. L. Smith, M. M. Bencivengo and C. A. Kunsch that indicated that prior growth of Staphylococcus aureus 196E on glycerol or maltose led to cells with repressed ability to produce staphylococcal enterotoxin A (J. L. Smith et al., "Entertoxin A Synthesis in Staphylococcus aureus: Inhibition by Glycerol and Maltose", Journal of General Microbiology, Vol 132, pp. 3375-3380, 1986).
Chloramphenicol has also been reported as totally inhibiting Staphylococcal enterotoxin B production (Robert A. Altenbern, "Protease Inhibitors Suppress Enterotoxin B Formation by Staphylococcus aureus", FEMS Microbiology Letters, Vol. 3, pp. 199-202, 1978).
The production of TSST-1 by S. aureus has predominantly been associated with menstrual toxic shock syndrome, which has in turn been related to tampon usage. Unexpectedly, a group of compounds were identified which are effective to substantially reduce toxic shock syndrome toxin 1 production by S. aureus in vitro and in vivo. These compounds and the method of their use in absorbent products are described in copending U.S. patent applications Ser. No. 343,965 filed Apr. 27, 1989 and Ser. No. 316,742 filed Apr. 27, 1990.
However, despite the effectiveness of the compounds in inhibiting the production of TSST-1 toxin, there is no basis for predicting whether such compounds would be effective against SEA, SEB, and SEC production due to the structural and chemical differences between the TSST-1 toxin and other Staphylococcal enterotoxins.